Structural basis for the modulation of MRP2 activity by phosphorylation and drugs
Abstract Multidrug resistance-associated protein 2 (MRP2/ABCC2) is a polyspecific efflux transporter of organic anions expressed in hepatocyte canalicular membranes. MRP2 dysfunction, in Dubin-Johnson syndrome or by off-target inhibition, for example by the uricosuric drug probenecid, elevates circu...
| Main Authors: | , , , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Nature Portfolio
2024-03-01
|
| Series: | Nature Communications |
| Online Access: | https://doi.org/10.1038/s41467-024-46392-8 |
| _version_ | 1797274000895770624 |
|---|---|
| author | Tiziano Mazza Theodoros I. Roumeliotis Elena Garitta David Drew S. Tamir Rashid Cesare Indiveri Jyoti S. Choudhary Kenneth J. Linton Konstantinos Beis |
| author_facet | Tiziano Mazza Theodoros I. Roumeliotis Elena Garitta David Drew S. Tamir Rashid Cesare Indiveri Jyoti S. Choudhary Kenneth J. Linton Konstantinos Beis |
| author_sort | Tiziano Mazza |
| collection | DOAJ |
| description | Abstract Multidrug resistance-associated protein 2 (MRP2/ABCC2) is a polyspecific efflux transporter of organic anions expressed in hepatocyte canalicular membranes. MRP2 dysfunction, in Dubin-Johnson syndrome or by off-target inhibition, for example by the uricosuric drug probenecid, elevates circulating bilirubin glucuronide and is a cause of jaundice. Here, we determine the cryo-EM structure of rat Mrp2 (rMrp2) in an autoinhibited state and in complex with probenecid. The autoinhibited state exhibits an unusual conformation for this class of transporter in which the regulatory domain is folded within the transmembrane domain cavity. In vitro phosphorylation, mass spectrometry and transport assays show that phosphorylation of the regulatory domain relieves this autoinhibition and enhances rMrp2 transport activity. The in vitro data is confirmed in human hepatocyte-like cells, in which inhibition of endogenous kinases also reduces human MRP2 transport activity. The drug-bound state reveals two probenecid binding sites that suggest a dynamic interplay with autoinhibition. Mapping of the Dubin-Johnson mutations onto the rodent structure indicates that many may interfere with the transition between conformational states. |
| first_indexed | 2024-03-07T14:53:06Z |
| format | Article |
| id | doaj.art-fdd5bced4bd2462399776434cd49d9e5 |
| institution | Directory Open Access Journal |
| issn | 2041-1723 |
| language | English |
| last_indexed | 2024-03-07T14:53:06Z |
| publishDate | 2024-03-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Nature Communications |
| spelling | doaj.art-fdd5bced4bd2462399776434cd49d9e52024-03-05T19:38:24ZengNature PortfolioNature Communications2041-17232024-03-0115111410.1038/s41467-024-46392-8Structural basis for the modulation of MRP2 activity by phosphorylation and drugsTiziano Mazza0Theodoros I. Roumeliotis1Elena Garitta2David Drew3S. Tamir Rashid4Cesare Indiveri5Jyoti S. Choudhary6Kenneth J. Linton7Konstantinos Beis8Department of Life Sciences, Imperial College LondonFunctional Proteomics group, Chester Beatty Laboratories, The Institute of Cancer ResearchBlizard Institute, Faculty of Medicine and Dentistry, Queen Mary University of LondonDepartment of Biochemistry and Biophysics, Stockholm UniversityDepartment of Metabolism, Digestion & Reproduction, Imperial College LondonDepartment DiBEST (Biologia, Ecologia, Scienze Della Terra) Unit of Biochemistry and Molecular Biotechnology, University of CalabriaFunctional Proteomics group, Chester Beatty Laboratories, The Institute of Cancer ResearchBlizard Institute, Faculty of Medicine and Dentistry, Queen Mary University of LondonDepartment of Life Sciences, Imperial College LondonAbstract Multidrug resistance-associated protein 2 (MRP2/ABCC2) is a polyspecific efflux transporter of organic anions expressed in hepatocyte canalicular membranes. MRP2 dysfunction, in Dubin-Johnson syndrome or by off-target inhibition, for example by the uricosuric drug probenecid, elevates circulating bilirubin glucuronide and is a cause of jaundice. Here, we determine the cryo-EM structure of rat Mrp2 (rMrp2) in an autoinhibited state and in complex with probenecid. The autoinhibited state exhibits an unusual conformation for this class of transporter in which the regulatory domain is folded within the transmembrane domain cavity. In vitro phosphorylation, mass spectrometry and transport assays show that phosphorylation of the regulatory domain relieves this autoinhibition and enhances rMrp2 transport activity. The in vitro data is confirmed in human hepatocyte-like cells, in which inhibition of endogenous kinases also reduces human MRP2 transport activity. The drug-bound state reveals two probenecid binding sites that suggest a dynamic interplay with autoinhibition. Mapping of the Dubin-Johnson mutations onto the rodent structure indicates that many may interfere with the transition between conformational states.https://doi.org/10.1038/s41467-024-46392-8 |
| spellingShingle | Tiziano Mazza Theodoros I. Roumeliotis Elena Garitta David Drew S. Tamir Rashid Cesare Indiveri Jyoti S. Choudhary Kenneth J. Linton Konstantinos Beis Structural basis for the modulation of MRP2 activity by phosphorylation and drugs Nature Communications |
| title | Structural basis for the modulation of MRP2 activity by phosphorylation and drugs |
| title_full | Structural basis for the modulation of MRP2 activity by phosphorylation and drugs |
| title_fullStr | Structural basis for the modulation of MRP2 activity by phosphorylation and drugs |
| title_full_unstemmed | Structural basis for the modulation of MRP2 activity by phosphorylation and drugs |
| title_short | Structural basis for the modulation of MRP2 activity by phosphorylation and drugs |
| title_sort | structural basis for the modulation of mrp2 activity by phosphorylation and drugs |
| url | https://doi.org/10.1038/s41467-024-46392-8 |
| work_keys_str_mv | AT tizianomazza structuralbasisforthemodulationofmrp2activitybyphosphorylationanddrugs AT theodorosiroumeliotis structuralbasisforthemodulationofmrp2activitybyphosphorylationanddrugs AT elenagaritta structuralbasisforthemodulationofmrp2activitybyphosphorylationanddrugs AT daviddrew structuralbasisforthemodulationofmrp2activitybyphosphorylationanddrugs AT stamirrashid structuralbasisforthemodulationofmrp2activitybyphosphorylationanddrugs AT cesareindiveri structuralbasisforthemodulationofmrp2activitybyphosphorylationanddrugs AT jyotischoudhary structuralbasisforthemodulationofmrp2activitybyphosphorylationanddrugs AT kennethjlinton structuralbasisforthemodulationofmrp2activitybyphosphorylationanddrugs AT konstantinosbeis structuralbasisforthemodulationofmrp2activitybyphosphorylationanddrugs |